In this study special emphasis is placed on metalloproteinase activity during tumor-endothelial cell interaction. Endothelial cDNA library was used to isolate clones for mammalian collagenase and tissue inhibitor of metalloproteinase (TIMP), both of which were found to be identical to the published sequences for the fibroblast collagenase and TIMP genes. Tumor cells exposed to medium from endothelial cells expressed a reduction in collagenolytic activity. Exposure of microvascular endothelial cells to tumor cell conditioned medium resulted in increased TIMP expression in contrast to normal cell conditioned medium which had no effect. TGFbeta treatment reduced TIMP expression of endothelial cells, fibroblasts, and tumor cell lines. A syngeneic rat model used to study micrometastasis consisted of raf/myc transfected rat liver epithelial cells carrying the E. coli lacZ gene as a reporter gene. Following intravenous injection into syngeneic rats these highly malignant cells could be visualized by histochemical staining for Beta-D-galactosidase. Individual cells were detected in lungs within 15 minutes and micrometastases after five days. Histopathological study of intravascular tumor thrombi was performed on disseminating hepatocellular carcinoma in nonhuman primates. Aldehyde fuchsin staining revealed that basement membrane was not disrupted, although the vascular lumen was distended with tumor thrombus. The thrombi were frequently covered with a single layer of endothelial cells. These findings indicate that in vivo the tumors may attach and grow intravascularly and be endothelialized before they break through the vascular basement membrane and grow as metastases. To continue this study we will specifically focus on metalloproteinase and TIMP expression in tissue sections showing vascular invasion, using the in situ hybridization technique.